This study was to compare the effects of parenteral supplementation of methionyl-methionine (Met-Met) or Met on intestinal barrier function in Met-deficient pregnant mice. Pregnant mice were randomly divided into three groups. The Control group was provided a diet containing Met and received i.p. injection of saline. The Met group was fed the same diet but without Met and received daily i.p. injection of 35% of the Met contained in the control diet. The Met-Met group was treated the same as the Met group, except that 25% of the Met injected was replaced with Met-Met. Met-Met promoted villus surface area in ileum compared with Met alone. In addition, the mRNA abundance of amino acid and glucose transporters in the small intestine was altered with Met-Met. Moreover, Met-Met increased tight junction protein and decreased apoptosis-related proteins expression in the jejunum and ileum. These results suggest that Met-Met can promote intestinal function over Met alone in Met-deficient mice.